The premise of this R21 is to translate the rodent work in the Chiu laboratory to a gene therapy for progressive multiple sclerosis (MS). MS is the major demyelinating disease for neurological disability in young adults in the US. MS starts with an early phase but converts into a late or progressive phase that is currently incurable. From 2015-2019, the Chiu laboratory identified a specific mechanism for neurodegeneration in progressive MS by studying the progressive phase of two different mouse models (Shiverer and EAE/NOD) that mimic the complex range of human MS. The culprit is an axon-specific mitochondrial anchoring protein called syntaphilin (SNPH), which is excessively upregulated as the disease transitions into the late phase. The result is excessive braking of mitochondria along the axons that disrupts recycling of mitochondria, leading to degradation in mitochondrial health that ultimately kills the axons. In the last 4 years, the Chiu laboratory demonstrated that the two models of progressive MS can be rescued by constitutive SNPH-KO intervention. The goal of this R21 is to see if similar rescue can be achieved by CRISPR-AAV gene therapy. We have designed CRISPR molecular scissors to cut-and-silence SNPH and will package the scissors into novel AAV vectors that can penetrate the blood-brain-barrier by single-dose intravenous injections. In Aim #1, we will use a CNS vector (AAV-PHP.eB) to treat the Shiverer mouse to rescue the cerebellum. In Aim #2, we will use a PNS vector (AAV-PHP.S) to treat the EAE/NOD mouse to rescue the spinal cord by promoting remyelination. Conclusion ? There are many FDA-approved drugs for the early phase of MS, but no effective treatments are available once the disease progresses into the late phase. In the last 4 years, the Chiu laboratory has identified excessive braking of mitochondria as a cause for killing axons in the progressive phase using two rodent models. This R21 grant will translate 4-years of rodent work into a possible gene therapy to treat human progressive MS.